Abstract
Hypermethylation of promoter CpG islands is generally recognized epigenetic mechanism responsible for gene silencing in cancer. However, molecular details on how this epigenetic mark triggers the process of gene downregulation are still elusive. Here, we used deep bisulfite sequencing and qPCR analysis to investigate the pattern of CpG methylation of ALDH1L1 promoter region and its association with the gene expression level in 16 paired breast cancer (BC) samples of different clinical stages. Expression of ALDH1L1 gene was suppressed in all examined BC samples up to 200-fold, and average hypermethylation level of the promoter region correlated positively with ALDH1L1 downregulation. We determined the role of every individual CpG site within the ALDH1L1 promoter, including upstream untranscribed region, first untranslated exon, and the start of the first intron, in aberrant gene expression by correlation analysis. The search revealed CpG sites which methylation has the highest impact on intensity of gene transcription. The majority of such CpG sites are located in a compact region in the first intron of the ALDH1L1 gene. These results assist in unraveling of dynamic nature of CpG promoter hypermethylation as well as demonstrate the efficiency of deep bisulfite sequencing in search for novel epigenetic markers in cancer.
Highlights
Altered protein expression pattern distinguishes cancer cells from normal ones
We have demonstrated that ALDH1L1 gene was strongly suppressed in 100% of breast cancer (BC) samples
Our analysis of TCGA data showed that suppression of the gene in BC is accompanied by CpG hypermethylation of its promoter region
Summary
Altered protein expression pattern distinguishes cancer cells from normal ones. These changes concern tumor suppressors or oncogene proteins but a variety of cellular proteins and believed to provide a selective advantage for uncontrolled proliferation, one of the hallmarks of cancer (Hanahan and Weinberg, 2000; Hanash and Taguchi, 2010). ALDH1L1 (aldehyde dehydrogenase 1 family, member L1) is one of the genes which expression is strongly downregulated in many human cancers including hepatocellular carcinoma, pilocytic astrocytoma, liver cancer (Krupenko and Oleinik, 2002; Rodriguez et al, 2008; Chen et al, 2012), renal cell carcinoma (Dmitriev et al, 2014), lung adenocarcinoma (Oleinik et al, 2011). The product of the ALDH1L1 gene, 10formyltetrahydrofolate dehydrogenase (FDH), is an abundant cytosolic enzyme involved in folate pathways (Krupenko, 2009). The observation that FDH is a strong marker of astrocytes in the rat brain suggests a function for the enzyme in the nervous system (Cahoy et al, 2008)
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